Chlorophyll a concentration (Chla) is a well-proven proxy of biocrust development, photosynthetic organisms\u2019 status, and recovery monitoring after environmental disturbances. However, laboratory methods for the analysis of chlorophyll require destructive sampling and are expensive and time consuming. Indirect estimation of chlorophyll a by means of soil surface reflectance analysis has been demonstrated to be an accurate, cheap, and quick alternative for chlorophyll retrieval information, especially in plants. However, its application to biocrusts has yet to be harnessed. In this study we evaluated the potential of soil surface reflectance measurements for non-destructive Chla quantification over a range of biocrust types and soils. Our results revealed that from the different spectral transformation methods and techniques, the first derivative of the reflectance and the continuum removal were the most accurate for Chla retrieval. Normalized difference values in the red-edge region and common broadband indexes (e.g., normalized difference vegetation index (NDVI)) were also sensitive to changes in Chla. However, such approaches should be carefully adapted to each specific biocrust type. On the other hand, the combination of spectral measurements with non-linear random forest (RF) models provided very good fits (R2 > 0.94) with a mean root mean square error (RMSE) of about 6.5 \u00b5g/g soil, and alleviated the need for a specific calibration for each crust type, opening a wide range of opportunities to advance our knowledge of biocrust responses to ongoing global change and degradation processes from anthropogenic disturbance.